We intend to study in vitro the mechanism by which the binding of cholinergic agonists to the nicotinic cholinergic receptor causes a change of membrane permeability of the post-synaptic membrane. Post- synaptic membranes will be isolated from Torpedo electric tissue, and fluorescence and radioisotope techniques will be used to characterize the equilibrium binding of cholinergic ligands and the steady-state membrane response. The correlation between receptor affinity and the permeability response will be utilized to identify receptor conformations associated with active and desensitized receptors. The actiion of drugs such as local anesthetics and other non-competitive antagonists will be studied to determine the mechanism by which they modify the cholinergic respones. Detailed studies will be undertaken to characterize the site of action of local anesthetics in terms of the membrane components to which they bind and the relationship to the receptor binding site. An analysis will be made of the functional significance of the different components (protein, lipid, carbohydrate) of the post-synaptic membrane by studying the effect on the cholinergic response of controlled enzymatic and chemical modifications. The membranes will be fractionated to identify components other than the receptor protein that are important for the control of receptor affinity and the permeability response. Fluorescence techniques will be used to identify structural changes of the membrane associated with the permeability response and to study the role of protein-protein and protein-lipid interactions in these membranes. Techniques will be developed to isolate and characterize more complete synaptic junctional complexes and to isolate post-synaptic membranes from tissue other than Torpedo electric tissue.